Elucidation of the intracellular mechanisms of cisplatin-induced apoptosis in renal cell line

本实验研究了顺铂诱导温度敏感猴病毒40抗原基因转基因小鼠终末近端直小管（S_3）细胞凋亡的细胞内机制。(1)caspase的参与：通过使用肽类caspase抑制剂和引入编码caspase抑制剂的病毒基因，发现caspase-3参与顺铂诱导的细胞凋亡，而一些caspase-3以外的caspase参与蛋白激酶C抑制剂staurosporine诱导的细胞凋亡。因此，不同的caspase可能参与了不同刺激诱导的S_3细胞凋亡。通过RT-PCR分析小鼠单个肾单位片段的mRNA，检测caspase-3在肾单位内的分布。Caspase-3 mRNA在小鼠肾单位各节段均有表达，以远曲小管和集合管表达最高。(2)与细胞周期进程0的相关性：caspase抑制可增加G2期的程度， ...更多信息 结果表明，caspase的激活可能通过减少阻滞在G2期的细胞数量而诱导S_3细胞凋亡。(3)氧自由基的参与：用荧光探针和共聚焦激光扫描显微镜观察顺铂处理S_3细胞后H_2O_2的产生。过氧化氢酶抑制顺铂诱导的S_3细胞H_2O_2的产生和细胞毒作用。此外，H_2O_2还可诱导S_3细胞凋亡、坏死、胀亡和凋亡性胀亡。H_2O_2处理的S_3细胞中存在DNA片段化现象，说明DNA片段化并非细胞凋亡所特有。(4)癌基因的参与：顺铂处理的S_3细胞c-fos mHNA表达增加。bcl-2过表达抑制caspase-3的激活和顺铂诱导的S_3细胞凋亡。这些结果提示caspase、细胞周期进程、自由基氧自由基和癌基因参与了顺铂诱导的S_3细胞凋亡。少

We have examined the intracellular mechanisms of cisplatin-induced apoptosis in mouse terminal proximal straight tubule (S_3) cell line derived from transgenic mice harboring temperature-sensitive simian virus 40 antigen gene. (1) The involvement of caspase : Using the peptide caspase inhibitor and the introduction of viral gene encoding caspase inhibitor, it appears that caspase-3 is involved in cisplatin-induced apoptosis, whereas some caspases other than caspase-3 in apoptosis induced by staurosporine , protein kinase C inhibitor. Thus, different caspases may be involved in apoptosis in S_3 cells induced by different stimuli. The intranephron distribution of caspase-3 was examined by RT-PCR analysis from mRNA of mouse individual nephron segments. Caspase-3 mRNA was expressed in all of mouse nephron segments tested, and those for distal tubule and collecting duct were the highest. (2) The association with cell cycleprogression 0 : The inhibition of caspases increased the degree of G2 … More arrest in cisplatin-treated S_3 cells, suggesting that the activation of caspases induces apoptosis by decreasing the number of cells arrested at G2 phase of the cell cycle in cisplatin-treated S_3 cells. (3) The involvement of free radical oxygene species : Using fluorescent probe and cofocal laser scanning microscope, H_2O_2 production was detected in cisplatin-treated S_3 cells. The treatment with catalase inhibited H_2O_2 production and the cytotoxicity in cisplatin-induced S_3 cells. In addition, H_2O_2 was shown to induce apoptosis, necrosis, oncosis and apoptotic oncosis in S_3cells. Since DNA fragmentation was detected in H_2O_2-treated S_3 cells, DNA fragmentation appeared not to be unique to apoptosis. (4) The involvement of oncogene : The increase in c-fos mHNA expression was observed in cisplatin-treated S_3 cells. Overexpression of bcl-2 inhibited the activation of caspase-3 and cell death in cisplatin-treated S_3 cellsThese results suggest that caspase, cell cycle progression, free radical oxygen species and oncogene are involved in cisplatin-induced apoptosis in S_3 cells. Less